Since its conception more than 27 years ago, the principle of particle counting and sizing invented by Wallace H. Coulter has resulted in numerous methods and apparatuses for the electronic counting, sizing and analysis of microscopic particles, which are scanned in a fluid suspension, as shown by the pioneer U.S. Pat. No. 2,656,508 to Coulter. In this prior art arrangement, a D.C. electric current flow is established between two vessels by suspending electrodes in the respective bodies of the suspension fluid. The only fluid connection between the two bodies is through a sensing orifice; hence, an electric current flow and field are established in the orifice. The orifice and the resultant electric field in and around it constitute a sensing zone. As each particle passes through the sensing zone, for the duration of the passage, the impedance of the contents of the sensing zone will change, thereby modulating the current flow and electric field in the sensing zone, and hence causing the generation of a signal to be applied to a detector suitably arranged to respond to such change. (The mark "Coulter" is a registered trademark, Registration No. 995,825, of Coulter Electronics, Inc. of Hialeah, Florida.)
To date, sensing orifices have been made in such a way that their entrances and exits were rather abrupt, so as to minimize the rise time associated with any particular particle's signal. From a fluid dynamics standpoint, orifices with sharp edges, or even rounded edges, are undesirable.
For a classical aperture opening into a semi-infinite space, for flow into the aperture, it is fairly well known that the flow pattern is such that the streamlines are roughly radial and the isobars are roughly hemispherical. In other words, the flow field is not at all similar to the same aperture system with flow from the aperture into the quiescent fluid space.
The important effect of the nearly hemispherical isobars is that the velocity gradient experienced by a particle drawn into the aperture is quite steep. In practice, large and/or dense particles simply do not accelerate as quickly as does the suspending fluid. This is especially true of the high Reynolds numbers (&gt;&gt;1) associated with apertures much larger than 100 .mu.m in diameter. The significance of the substantial Reynolds number is that the inertia forces acting on the fluid and the particle become relatively more important than the fluid shear forces acting on fluid or particle, with increasing Reynolds number.
It has been experimentally verified that large and/or dense particles traverse a sharp edged aperture at velocities substantially lower than the peak fluid velocity, or even the bulk fluid velocity. The slow pulse rise times associated with these slow-moving particles can cause difficulties in the signal processing electronics, and the long pulse lengths can cause problems with coincidence. Attempts to increase the particle velocity in the aperture by increasing the aperture flow rate have met with rapidly diminishing returns, since the pressure drop required increases rapidly, and there is a greater velocity shortfall for higher flow rates.
It should be noted that the highly esteemed "hydrodynamic focusing" technique that works so well at low Reynolds numbers becomes a dismal failure at high Reynolds numbers, because of the steep velocity gradient associated with the "focusing" zone. This ties together some problems associated with an aperture's entrance.
There is also a problem associated with the aperture's exit. The velocity distribution associated with a stream of fluid leaving an aperture and entering a quiescent pool of similar or identical fluid is generally pencil-like. If the receiving chamber is smaller than semi-infinite, the presence of a highly directional jet entering the chamber will induce a generally toroidal circulation within the chamber. In a particle counter of the electrical sensing zone type, it is known that the sensing zone extends beyond both ends of the aperture (symmetrically, unlike the fluid flow fields), has isopotential surfaces which are roughly hemispherical, and can detect the presence of a particle without the aperture and removed by an aperture diameter or more. The toroidal recirculation will eventually carry particles which have already been counted back into the downstream area of the sensing zone, causing them to be counted again and again. A technique called "sweep flow" can prevent this recirculation, but is not likely to be fully effective at high Reynolds numbers, requires complex plumbing and does not diminish the size of the downstream sensing zone.
Accordingly, it can be seen that there is a need in the art for a flow cell which attacks and overcomes all of the problems just discussed and reiterated as follows: (A) velocity shortfall due to steep gradients in the aperture entrance, causing slow-rising pulses and donsequent distortion of small signals by AC-coupled electronics; (B) recirculation of particles behind the aperture, causing multiple false counts; and (C) electrical sensing zone substantially longer than the aperture's physical length, aggravating so-called vertical coincidence and making it difficult to circumvent the effects of same Velocity shortfall also aggravates this problem.
Swedish Pat. No. 355,959 to Everaerts discloses a capacitor formed from two concentric, metal, ring-like electrodes surrounding a moving liquid sample, which are energized with a high frequency source to determine the conductivity of the sample by measuring its capacitance and resistance in a resonating circuit. A typical "focused flow" in which particles are hydrodynamically focused as they pass through a sensing orifice is shown in the article entitled "A Volume-Activated Cell Sorter", THE JOURNAL OF HISTOCHEMISTRY AND CYTOCHEMISTRY, Menke et al, Vol. 25, No. 7, pp. 796-803, 1977.
East German Pat. No. 66,038 a plurality of electrodes mounted along the entire length of an aperture for sensing particles in multiple sensing zones.
U.S. Pat. No. 4,140,966 to Godin et al is incorporated herein.